Chlamydia trachomatis infection is the leading cause of infertility and ectopic pregnancy in women. Infection is often asymptomatic, leading to lack of detection, increased risk for disease and delay in treatment. Although antibiotic therapy eliminates infection, it does not mitigate or reverse established destructive pathology. Thus, there is a critical need for an effective preventative approach, in the form of a vaccine. Rational vaccine design requires understanding of the adaptive mechanisms associated with protective immunity and identification of chlamydial antigens that best elicit protective responses. Animal models and human epidemiologic data indicate that repeated exposure will ultimately protect against reinfection. Resolution of infection by attenuated chlamydial mutants protects against genital and ocular disease, providing evidence that an effective chlamydial vaccine can be developed. Mouse model and human data indicate that IFN-?-producing, Chlamydia-specific CD4 (Th1) cells are the primary mediators of protective immunity. We have identified chlamydial proteins that elicit IFN-? production predominantly from CD4 T cells and are significantly associated with protection against chlamydial reinfection in a group of highly sexually active women. Transcriptional profiling of women with chlamydial pelvic inflammatory disease reveal dysregulation of genes involved in T cell growth, proliferation, and signaling. These data support our central hypothesis that protection is dependent on the generation of antigen-specific CD4 T cell responses. Our overall objectives are to validate our novel vaccine candidate antigens, profile the T cells recognizing these antigens, and determine local and systemic responses that lead to an effective anti- chlamydial immune response. Our rationale is that identification of T cell effector activities and antigens associated with protection will advance the development of a vaccine. We will test our central hypothesis by completing the following specific aims: 1. Characterize CD4 and CD8 T cells recognizing chlamydial antigens associated with resistance to reinfection. We will test the hypothesis that polyfunctional central and effector memory CD4 but not CD8 Th1 cells are induced in protected women, using multiparameter flow cytometry, multiplex cytokine and mRNA analyses. 2. Define a transcriptional signature that leads to protection from reinfection. We will use endometrial RNAseq, whole blood and cellular microarrays to elucidate the evolution of a protective adaptive immune response to Chlamydia and to identify surrogate markers of vaccine efficacy. Expected outcomes of this work are important information related to vaccine antigens, T cell memory responses, and signaling pathways that correlate with protection against the world's most prevalent sexually transmitted bacterial pathogen and a leading cause of preventable blindness.